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Kinetics and yields of pesticide biodegradation at low substrate concentrations and under conditions restricting assimilable organic carbon.

ABSTRACT: The fundamentals of growth-linked biodegradation occurring at low substrate concentrations are poorly understood. Substrate utilization kinetics and microbial growth yields are two critically important process parameters that can be influenced by low substrate concentrations. Standard biodegradation tests aimed at measuring these parameters generally ignore the ubiquitous occurrence of assimilable organic carbon (AOC) in experimental systems which can be present at concentrations exceeding the concentration of the target substrate. The occurrence of AOC effectively makes biodegradation assays conducted at low substrate concentrations mixed-substrate assays, which can have profound effects on observed substrate utilization kinetics and microbial growth yields. In this work, we introduce a novel methodology for investigating biodegradation at low concentrations by restricting AOC in our experiments. We modified an existing method designed to measure trace concentrations of AOC in water samples and applied it to systems in which pure bacterial strains were growing on pesticide substrates between 0.01 and 50 mg liter(-1). We simultaneously measured substrate concentrations by means of high-performance liquid chromatography with UV detection (HPLC-UV) or mass spectrometry (MS) and cell densities by means of flow cytometry. Our data demonstrate that substrate utilization kinetic parameters estimated from high-concentration experiments can be used to predict substrate utilization at low concentrations under AOC-restricted conditions. Further, restricting AOC in our experiments enabled accurate and direct measurement of microbial growth yields at environmentally relevant concentrations for the first time. These are critical measurements for evaluating the degradation potential of natural or engineered remediation systems. Our work provides novel insights into the kinetics of biodegradation processes and growth yields at low substrate concentrations.

SUBMITTER: Helbling DE

PROVIDER: S-EPMC3911073 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Kinetics and yields of pesticide biodegradation at low substrate concentrations and under conditions restricting assimilable organic carbon.

Helbling Damian E DE Hammes Frederik F Egli Thomas T Kohler Hans-Peter E HP

Applied and environmental microbiology 20131206 4

The fundamentals of growth-linked biodegradation occurring at low substrate concentrations are poorly understood. Substrate utilization kinetics and microbial growth yields are two critically important process parameters that can be influenced by low substrate concentrations. Standard biodegradation tests aimed at measuring these parameters generally ignore the ubiquitous occurrence of assimilable organic carbon (AOC) in experimental systems which can be present at concentrations exceeding the c ...[more]

PMID: 24317077

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Project description:The objective of our study was to test whether limited microbial degradation at low pesticide concentrations could explain the discrepancy between overall degradability demonstrated in laboratory tests and their actual persistence in the environment. Studies on pesticide degradation are often performed using unrealistically high application rates seldom found in natural environments. Nevertheless, biodegradation rates determined for higher pesticide doses cannot necessarily be extrapolated to lower concentrations. In this context, we wanted to (i) compare the kinetics of pesticide degradation at different concentrations in arable land and (ii) clarify whether there is a concentration threshold below which the expression of the functional genes involved in the degradation pathway is inhibited without further pesticide degradation taking place. We set up an incubation experiment for four weeks using 14C-ring labeled 2-methyl-4-chlorophenoxyacetic acid (MCPA) as a model compound in concentrations from 30 to 20,000 μg kg-1 soil. To quantify the abundance of putative microorganisms involved in MCPA degradation and their degradation activity, tfdA gene copy numbers (DNA) and transcripts (mRNA) were determined by quantitative real-time PCR. Mineralization dynamics of MCPA derived-C were analyzed by monitoring 14CO2 production and 14C assimilation by soil microorganisms. We identified two different concentration thresholds for growth and activity with respect to MCPA degradation using tfdA gene and mRNA transcript abundance as growth and activity indices, respectively. The tfdA gene expression started to increase between 1,000 and 5,000 μg MCPA kg-1 dry soil, but an actual increase in tfdA sequences could only be determined at a concentration of 20,000 μg. Accordingly, we observed a clear shift from catabolic to anabolic utilization of MCPA-derived C in the concentration range of 1,000 to 5,000 μg kg-1. Concentrations ≥1,000 μg kg-1 were mainly associated with delayed mineralization, while concentrations ≤1,000 μg kg-1 showed rapid absolute dissipation. The persistence of pesticides at low concentrations cannot, therefore, be explained by the absence of functional gene expression. Nevertheless, significant differences in the degradation kinetics of MCPA between low and high pesticide concentrations illustrate the need for studies investigating pesticide degradation at environmentally relevant concentrations.

Project description:BACKGROUND: The expression and activity of the different Saccharomyces cerevisiae hexose uptake systems (Hxt) and the kinetics of glucose uptake are considered essential to industrial alcoholic fermentation performance. However, the dynamics of glucose uptake kinetics during the different stages of fermentation, depending on glucose and nitrogen availability, is very poorly characterized. The objective of the present work was to examine thoroughly the alterations occurring in glucose uptake kinetics during alcoholic fermentation, by the wine strain S. cerevisiae PYCC 4072, of a synthetic grape juice basal medium with either a limiting or non-limiting initial nitrogen concentration and following nitrogen supplementation of the nitrogen-depleted sluggish fermentation. RESULTS: Independently of the initial concentration of the nitrogen source, glucose transport capacity is maximal during the early stages of fermentation and presumably sustained by the low-affinity and high-capacity glucose transporter Hxt1p. During nitrogen-limited sluggish fermentation, glucose uptake capacity was reduced to approximately 20% of its initial values (Vmax?=?4.9?±?0.8 compared to 21.9?±?1.2??mol?h?¹ 10?? cells), being presumably sustained by the low-affinity glucose transporter Hxt3p (considering the calculated Km?=?39.2?±?8.6?mM). The supplementation of the sluggish fermentation broth with ammonium led to the increase of glucose transport capacity associated to the expression of different glucose uptake systems with low and high affinities for glucose (Km?=?58.2?±?9.1 and 2.7?±?0.4?mM). A biclustering analysis carried out using microarray data, previously obtained for this yeast strain transcriptional response to equivalent fermentation conditions, indicates that the activation of the expression of genes encoding the glucose transporters Hxt2p (during the transition period to active fermentation) and Hxt3p, Hxt4p, Hxt6p and Hxt7p (during the period of active fermentation) may have a major role in the recovery of glucose uptake rate following ammonium supplementation. These results suggest a general derepression of the glucose-repressible HXT genes and are consistent with the downregulation of Mig1p and Rgt1p. CONCLUSIONS: Although reduced, glucose uptake rate during nitrogen-limited fermentation is not abrogated. Following ammonium supplementation, sluggish fermentation recovery is associated to the increase of glucose uptake capacity, related to the de novo synthesis of glucose transporters with different affinity for glucose and capacity, presumably of Hxt2p, Hxt3p, Hxt4p, Hxt6p and Hxt7p. This study is a contribution to the understanding of yeast response to different stages of alcoholic fermentation at the level of glucose uptake kinetics, in particular under nitrogen limitation or replenish, which is useful knowledge to guide fermentation practices.

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Kinetics and yields of pesticide biodegradation at low substrate concentrations and under conditions restricting assimilable organic carbon.

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Kinetics and yields of pesticide biodegradation at low substrate concentrations and under conditions restricting assimilable organic carbon.

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